Abstract
The gastrointestinal epithelial layer forms a physical and biochemical barrier that maintains the segregation between host and intestinal microbiota. The integrity of this barrier is critical in maintaining homeostasis in the body and its dysfunction is linked to a variety of illnesses, especially inflammatory bowel disease. Gut microbes, and particularly probiotic bacteria, modulate the barrier integrity by reducing gut permeability and reinforcing tight junctions. Probiotic Escherichia coli Nissle 1917 (EcN) is a good colonizer of the human gut with proven therapeutic efficacy in the remission of ulcerative colitis in humans. EcN positively modulates the intestinal epithelial barrier through upregulation and redistribution of the tight junction proteins ZO-1, ZO-2 and claudin-14. Upregulation of claudin-14 has been attributed to the secreted protein TcpC. Whether regulation of ZO-1 and ZO-2 is mediated by EcN secreted factors remains unknown. The aim of this study was to explore whether outer membrane vesicles (OMVs) released by EcN strengthen the epithelial barrier. This study includes other E. coli strains of human intestinal origin that contain the tcpC gene, such as ECOR63. Cell-free supernatants collected from the wild-type strains and from the derived tcpC mutants were fractionated into isolated OMVs and soluble secreted factors. The impact of these extracellular fractions on the epithelial barrier was evaluated by measuring transepithelial resistance and expression of several tight junction proteins in T-84 and Caco-2 polarized monolayers. Our results show that the strengthening activity of EcN and ECOR63 does not exclusively depend on TcpC. Both OMVs and soluble factors secreted by these strains promote upregulation of ZO-1 and claudin-14, and down-regulation of claudin-2. The OMVs-mediated effects are TcpC-independent. Soluble secreted TcpC contributes to the upregulation of ZO-1 and claudin-14, but this protein has no effect on the transcriptional regulation of claudin-2. Thus, in addition to OMVs and TcpC, other active factors released by these microbiota strains contribute to the reinforcement of the epithelial barrier.
Highlights
The gastrointestinal epithelial layer is the first line of defense against pathogens and the surface where the host interacts with microbiota
The positive effects of the probiotic Escherichia coli Nissle 1917 (EcN) on intestinal barrier function have been attributed to TcpC, a protein that mediates upregulation of the tight junctions. ZO (TJ) protein claudin-14 (Hering et al, 2014)
Regarding the effect of COF-SN, upregulation of ZO-1 and claudin-14 was almost abolished by TcpC deficiency in EcN, and significantly weakened in the case of ECOR63 tcpC (Figure 3A, dotted bars). These results show that TcpC contributes to the transcriptional regulation of ZO-1 and claudin-14, and suggest that other released factors may contribute to this regulation in ECOR63
Summary
The gastrointestinal epithelial layer is the first line of defense against pathogens and the surface where the host interacts with microbiota. This specialized epithelium forms a physical and biochemical barrier that maintains the segregation between host and intestinal microbiota. Several factors contribute to the epithelial barrier function, including the production of a mucin layer that covers the epithelial surface and prevents direct contact with intestinal microbes, the secretion of antimicrobial peptides, and the establishment of TJ between intestinal epithelial cells that seal the host tissue against the luminal environment. The TJ that connect adjacent intestinal epithelial cells are composed of different types of integral membrane proteins such as occludin, several claudins, tricellulin, and junctional adhesion molecules (Turner, 2009). Claudin-2, for instance, controls the movement of ions and enhances transepithelial water flux (Rosenthal et al, 2010)
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